PROJECT SUMMARY Developmental responses to nutrient stress reflect systems-level regulation -- the entire animal and its progeny can be affected. But how developmental physiology is coordinated across the animal and over generations is not well understood. The long-term goal of this project is to understand the molecular basis of persistent effects of nutrient stress in the roundworm C. elegans. The worm is an ideal model since it has evolved to thrive in feast and famine and its short generation time facilitates multigenerational studies. Preliminary results show that larval starvation causes germline tumors to develop during early adulthood (intragenerational effect), but that reduction of insulin/IGF signaling suppresses tumors by promoting ferroptosis. They also show that mater- nal dietary restriction protects progeny from starvation-induced tumors by reducing insulin/IGF signaling (inter- generational effect). Our studies also demonstrate epigenetic inheritance of increased starvation resistance and lifespan as well as altered gene expression following dauer arrest (transgenerational effect), and they sug- gest that small RNAs in the germ line mediate these effects. These preliminary results lay the foundation for mechanistic analysis of persistent effects of nutrient stress during development and across generations. The premise of this proposal is that early-life starvation compromises developmental integrity, but parental or an- cestral nutrient stress buffers progeny from starvation. The central hypothesis is that early-life starvation leads to development of adult germline tumors, but maternal provisioning and epigenetic inheritance protect progeny from such pathological effects of starvation. The objectives are to identify signaling and gene regulatory mech- anisms that mediate adaptation to nutrient stress across generations. The central hypothesis is supported by strong preliminary data as well as the literature. It will be tested with the following three aims: 1) Identify mech- anisms by which reduction of insulin/IGF signaling suppresses starvation-induced germline tumors, 2) Identify mechanisms by which maternal dietary restriction buffers progeny from pathological effects of early-life starva- tion, and 3) Identify regulatory mechanisms that mediate epigenetic inheritance of starvation resistance. Genet- ic, genomic, pharmacological and biochemical approaches will be used to complete these aims. This work is innovative for developing models that facilitate mechanistic analysis of intra-, inter- and transgenerational ef- fects of nutrient stress and for investigating ferroptosis as a tumor suppressor mechanism regulated by a FoxO transcription factor and insulin/IGF signaling. The contributions of the proposed work will be identification of regulatory mechanisms that mediate adaptation to nutrient stress across generations. These include mecha- nisms by which FoxO transcription factors suppress tumors as well as mechanisms for inheritance of starvation resistance. These contributions will be significant because they will fill critical gaps in understanding of how nutrient stress affects development, maternal provisioning, and epigenetic inheritance. The deeply conserved function of insulin/IGF signaling and FoxO factors suggests that mechanisms discovered will be conserved.